NODULE INCEPTION Recruits the Lateral Root Developmental Program for Symbiotic Nodule Organogenesis in Medicago truncatula

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Contributions of the hippocampus and medial prefrontal cortex to energy and body weight regulation

The effects of selective ibotenate lesions of the complete hippocampus (CHip), the hippocampal ventral pole (VP), or the medial prefrontal cortex (mPFC) in male rats were assessed on several measures related to energy regulation (i.e., body weight gain, food intake, body adiposity, metabolic activity, general behavioral activity, conditioned appetitive responding). The testing conditions were designed to minimize the nonspecific debilitating effects of these surgeries on intake and body weight. Rats with CHip and VP lesions exhibited significantly greater weight gain and food intake compared with controls. Furthermore, CHip-lesioned rats, but not rats with VP lesions, showed elevated metabolic activity, general activity in the dark phase of the light-dark cycle, and greater conditioned appetitive behavior, compared with control rats without these brain lesions. In contrast, rats with mPFC lesions were not different from controls on any of these measures. These results indicate that hippocampal damage interferes with energy and body weight regulation, perhaps by disrupting higher-order learning and memory processes that contribute to the control of appetitive and consummatory behavior.     

Association of MICA with rheumatoid arthritis independent of known HLA-DRB1 risk alleles in a family-based and a case control study

Introduction

The gene MICA encodes the protein major histocompatibility complex class I polypeptide-related sequence A. It is expressed in synovium of patients with rheumatoid arthritis (RA) and its implication in autoimmunity is discussed. We analyzed the association of genetic variants of MICA with susceptibility to RA.

Methods

Initially, 300 French Caucasian individuals belonging to 100 RA trio families were studied. An additional 100 independent RA trio families and a German Caucasian case-control cohort (90/182 individuals) were available for replication. As MICA is situated in proximity to known risk alleles of the HLA-DRB1 locus, our analysis accounted for linkage disequilibrium either by analyzing the subgroup consisting of parents not carrying HLA-DRB1 risk alleles with transmission disequilibrium test (TDT) or by implementing a regression model including all available data. Analysis included a microsatellite polymorphism (GCT)n and single-nucleotide polymorphisms (SNPs) rs3763288 and rs1051794.

Results

In contrast to the other investigated polymorphisms, the non-synonymously coding SNP MICA-250 (rs1051794, Lys196Glu) was strongly associated in the first family cohort (TDT: P = 0.014; regression model: odds ratio [OR] 0.46, 95% confidence interval [CI] 0.25 to 0.82, P = 0.007). Although the replication family sample showed only a trend, combined family data remained consistent with the hypothesis of MICA-250 association independent from shared epitope (SE) alleles (TDT: P = 0.027; regression model: OR 0.56, 95% CI 0.38 to 0.83, P = 0.003). We also replicated the protective association of MICA-250A within a German Caucasian cohort (OR 0.31, 95% CI 0.1 to 0.7, P = 0.005; regression model: OR 0.6, 95% CI 0.37 to 0.96, P = 0.032). We showed complete linkage disequilibrium of MICA-250 (D'' = 1, r²= 1) with the functional MICA variant rs1051792 (D'' = 1, r²= 1). As rs1051792 confers differential allelic affinity of MICA to the receptor NKG2D, this provides a possible functional explanation for the observed association.

Conclusions

We present evidence for linkage and association of MICA-250 (rs1051794) with RA independent of known HLA-DRB1 risk alleles, suggesting MICA as an RA susceptibility gene. However, more studies within other populations are necessary to prove the general relevance of this polymorphism for RA.     

Genetic diversity in Passiflora species determined by morphological and molecular characteristics

Morphological and molecular characteristics were studied in six wild species of Passiflora. There were statistically significant differences among these six species for all characteristics studied. Intra-specific variability was observed for number of flowers, number of fruits, number of seeds, fruit length, fruit width and leaf area. Cluster analysis using morphological data showed three groups: 1) P. palmeri var. sublanceolata, P. morifolia and P. foetida var. foetida, 2) P. coriacea and P. micropetala, and 3) P. suberosa. The dendrogram constructed using randomly amplified polymorphic DNA (RAPD) data showed six different groups for each species. The genetic distances among the 24 accessions ranged from 0.05 (between P. morifolia accessions P1 and P3) to 0.95 (P. coriacea accession 31 and P. palmeri var. sublanceolata accession 49). The species showed high morphological and molecular inter- and intraspecific variability.     

DNA binding in the central channel of bacteriophage T7 helicase-primase is a multistep process. Nucleotide hydrolysis is not required

Many helicases assemble into ring-shaped hexamers and bind DNA in their central channel. This raises the question as to how the DNA gets into the central channel to form a topologically linked complex. We have used the presteady-state stopped-flow kinetic method and protein fluorescence changes to investigate the mechanism of single-stranded DNA (ssDNA) binding to the bacteriophage T7 helicase-primase, gp4A'. We have found that the kinetics of 30-mer ssDNA binding to a preformed gp4A' hexamer in the presence of both Mg-dTMP-PCP and Mg-dTTP are similar, indicating that Mg-dTTP binding is sufficient and hydrolysis is not necessary for efficient DNA binding. Multiple transient changes in gp4A' fluorescence revealed a four-step mechanism for DNA binding with Mg-dTTP. These transient changes were analyzed by global fitting and kinetic simulation to determine the intrinsic rate constants of this four-step mechanism. The initial steps, including the bimolecular encounter of the DNA with the helicase and a subsequent conformational change, were fast. We propose that these initial steps of DNA binding occur at a readily accessible site, which is likely to be on the outside of the hexamer ring. The binding of the 30-mer ssDNA at this loading site is followed by slower conformational changes that allow the DNA to transit into the central channel of gp4A' via a ring-opening or threading pathway.     

A ring-opening mechanism for DNA binding in the central channel of the T7 helicase-primase protein

We have investigated the mechanism of binding single-stranded DNA (ssDNA) into the central channel of the ring-shaped T7 gp4A' helicase-primase hexamer. Presteady-state kinetic studies show a facilitated five-step mechanism and provide understanding of how a ring-shaped helicase can be loaded on the DNA during the initiation of replication. The effect of a primase recognition sequence on the observed kinetics suggests that binding to the helicase DNA-binding site is facilitated by transient binding to the primase DNA-binding site, which is proposed to be a loading site. The proposed model involves the fast initial binding of the DNA to the primase site on the outside of the helicase ring, a fast conformational change, a ring-opening step, migration of the DNA into the central channel of the helicase ring, and ring closure. Although an intermediate protein-DNA complex is kinetically stable, only the last species in the five-step mechanism is poised to function as a helicase at the unwinding junction.     

The α‐subunit of the trimeric GTPase Go2 regulates axonal growth

The Goα splice variants Go1α and Go2α are subunits of the most abundant G‐proteins in brain, Go1 and Go2. Only a few interacting partners binding to Go1α have been described so far and splice variant‐specific differences are not known. Using a yeast two‐hybrid screen with constitutively active Go2α as bait, we identified Rap1GTPase activating protein (Rap1GAP) and Girdin as interacting partners of Go2α, which was confirmed by co‐immunoprecipitation. Comparison of subcellular fractions from brains of wild type and Go2α?/? mice revealed no differences in the overall expression level of Girdin or Rap1GAP. However, we found higher amounts of active Rap1‐GTP in brains of Go2α deficient mutants, indicating that Go2α may increase Rap1GAP activity, thereby effecting the Rap1 activation/deactivation cycle. Rap1 has been shown to be involved in neurite outgrowth and given a Rap1GAP‐Go2α interaction, we found that the loss of Go2α affected axonal outgrowth. Axons of cultured cortical and hippocampal neurons prepared from embryonic Go2α?/? mice grew longer and developed more branches than those from wild‐type mice. Taken together, we provide evidence that Go2α regulates axonal outgrowth and branching.